Some aspects of the biogenesis of tRNA from primary gene transcripts to their final functional form in both E. coli and certain eucaryotes will be investigated. In particular, RNase P, essential for the processing of the 5' termini of tRNAs, will be purified and extensively characterized in terms of its interaction with precursor molecules. Studies of tRNA structure-function relationships will also be undertaken through an investigation of the nature of the defect in certain mutants of tRNA which do not produce sufficient quantities of gene transcripts for RNA sequence analysis. These will be studied by DNA sequence analysis of DNA restriction enzyme fragment containing the appropriate tRNA gene sequences. Sequence analysis of the RNA moiety (MIRNA) of wild type and mutant RNase P will be carried out. Both wild type and genetically altered E. coli tRNA precursor molecules and/or RNase P will be used respectively as substrates and enzyme in studies of RNase-tRNA precursor interactions. RNA nucleotide sequence analysis or Maxam-Gilbert DNA sequence analysis will be used to elucidate the nature of defects altering suppression or processing defects in mutants of tRNA Tyr. M1 DNA will be cloned in plasmid vectors. The appropriate DNA restriction enzyme fragment will then be isolated and its sequence determined.